Repopulation of rat liver by fetal hepatoblasts and adult hepatocytes transduced ex vivo with lentiviral vectors
Repopulation of rat liver by fetal hepatoblasts and adult hepatocytes transduced ex vivo with lentiviral vectors
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DOI：
10.1053/jhep.2003.50183
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发表时间：
2003-05-01
期刊：
影响因子：
13.5
通讯作者：
Shafritz, DA
中科院分区：
文献类型：
--
作者：
Oertel, M;Rosencrantz, R;Shafritz, DA
最近的研究表明，基于人类免疫缺陷病毒的慢病毒载体可以有效地在体外转导非分散的原代细胞，例如肝细胞。其他研究报告说，在某些条件下，肝脏可以用移植的肝细胞再现。在本研究中，我们结合了这些程序，通过用肝细胞和慢跑病毒载体转导记者基因，绿色荧光蛋白（GFP），用肝细胞和肝细胞重现大鼠肝脏来开发用于离体基因治疗的模型系统。当转基因由肝脏特异性白蛋白增强子/启动子驱动时，体内长期GFP表达（长达4个月），但是当使用巨细胞病毒（CMV）增强子/启动子时，会沉默。在反性/部分肝切除术的影响下，将移植的细胞大规模扩增（类似于10个细胞双倍），并且通过双重表达细胞移植标记，二肽基肽酶IV（DPPIV）和GFP来记录单个细胞中的再构和持续转基因表达。在该系统中，移植细胞的维持或扩展不取决于转基因的表达，因此确定不需要阳性选择以在多个移植，慢病毒转导的肝细胞的多个分裂之后保持转基因表达。 In conclusion, fetal hepatoblasts (liver stem/progenitor cells) can serve as efficient vehicles for e-v vivo gene therapy and suggest that liver-based genetic disorders that do not shorten hepatocyte longevity or cause liver damage, such as phenylketonuria, hyperbilirubinemias, familial hypercholesterolemia, primary oxalosis, and factor DC deficiency, among others, might be可以通过这种方法接受治疗。
Recent studies have shown that nondividing primary cells, such as hepatocytes, can be efficiently transduced in vitro by human immunodeficiency virus-based lentivirus vectors. Other studies have reported that, under certain conditions, the liver can be repopulated with transplanted hepatocytes. In the present study, we combined these procedures to develop a model system for ex vivo gene therapy by repopulating rat livers with hepatocytes and hepatoblasts transduced with a lentivirus vector expressing a reporter gene, green fluorescent protein (GFP). Long-term GFP expression in vivo (up to 4 months) was achieved when the transgene was driven by the liver-specific albumin enhancer/promoter but was silenced when the cytomegalovirus (CMV) enhancer/promoter was used. Transplanted cells were massively amplified (similar to 10 cell doublings) under the influence of retrorsine/partial hepatectomy, and both repopulation and continued transgene expression in individual cells were documented by dual expression of a cell transplantation marker, dipeptidyl peptidase IV (DPPIV), and GFP. In this system, maintenance or expansion of the transplanted cells did not depend on expression of the transgene, establishing that positive selection is not required to maintain transgene expression following multiple divisions of transplanted, lentivirus-transduced hepatic cells. In conclusion, fetal hepatoblasts (liver stem/progenitor cells) can serve as efficient vehicles for e-v vivo gene therapy and suggest that liver-based genetic disorders that do not shorten hepatocyte longevity or cause liver damage, such as phenylketonuria, hyperbilirubinemias, familial hypercholesterolemia, primary oxalosis, and factor DC deficiency, among others, might be amenable to treatment by this approach.